To better understand the antigenic nature and the immune response to P. carinii, we have undertaken to purify the major surface antigen of rat and human P. carinii. It is necessary to use P. carinii from both sources because antigenically they are different and specifically the major surface antigen in rat and human appear to be different. The major surface antigens of both rat and human P. carinii have been purified by treatment of intact organisms with lyticase followed by separation on a sizing column and subsequent separation by ion-exchange chromotography. Using the above protocol, we have purified both rat and human P. carinii antigens in sufficient quantities to do preliminary biochemical analysis. Using cross- linking studies, we have documented that although the molecular weight of the antigen is about 95- 100,000 on SDS-PAGE, in its natural form it appears to have a molecular weight of about 280,000-300,000. The antigen is a glycoprotein with about 10% of the composition of the size being accounted for by carbohydrates. We are presently attempting to isolate from a cDNA library a clone that encodes the gene for the surface antigen. Using antibodies against P. carinii, we have screened a cDNA library and obtained a number of reactive clones. Three of these clones have been sequenced, and are related but not identical, suggesting that multiple genes encode the major surface antigen. The identity of these clones was confirmed by obtaining amino acid sequence information from purified gp116. The sequence obtained was identical to the predicted amino acid sequence of the closed cDNA. We are in the process of further characterizing the genes encoding the surface antigen. We have also been able to demonstrate that P. carinii in short-term tissue cultures can incorporate S35-methionine into the major surface antigen. This then gives us a method for evaluating the metabolism of teris protein. The goal of this study is to better understand the pathogenesis of P. carinii pneumonia, with the hope that we can use this information to control or prevent this disease.